CN108102924A - A kind of method of germ contamination in control chlorella culture using bacteriophagic Bdellovibrio - Google Patents
A kind of method of germ contamination in control chlorella culture using bacteriophagic Bdellovibrio Download PDFInfo
- Publication number
- CN108102924A CN108102924A CN201810030528.1A CN201810030528A CN108102924A CN 108102924 A CN108102924 A CN 108102924A CN 201810030528 A CN201810030528 A CN 201810030528A CN 108102924 A CN108102924 A CN 108102924A
- Authority
- CN
- China
- Prior art keywords
- chlorella
- culture
- bacteriophagic bdellovibrio
- bdellovibrio
- bacteriophagic
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/12—Unicellular algae; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
The present invention relates to one kind in chlorella is cultivated, it is infected by adding in bacteriophagic Bdellovibrio and kills the bacteriums of other pollutions, and then the method for improving chlorella growth, there is important application value in terms of the germ contamination control and raising chlorella growth in chlorella incubation.
Description
Technical field
The invention belongs to biological technical fields, are related to a kind of utilization bacteriophagic Bdellovibrio (Bdellovibrio
Bacteriovorus sp.) other germ contaminations in chlorella (Chlorella sp.) culture are controlled, and then promote chlorella
The method of growth,
Background technology
As one of simplest photosynthesis organism, chlorella is the microalgae that the mankind can cultivate earliest, not only
Growth is fast, and is rich in a variety of nutrition and the bioactive ingredients such as protein, polysaccharide and vitamin and chlorella growth factor,
It is subject to common concern both domestic and external, is widely used in human health care's food, bioenergy and environmental improvement etc..
In general chlorella culture has 3 kinds of illumination autotrophy, mixed breeding and no light Heterotrophic culture modes, but either tests
Room or industrialization large-scale culture are inevitably present different degrees of germ contamination situation.Because bacterium can by with
The modes such as chlorella competition nutrition, secretion anti-algal substance and dissolving frustule inhibit the growth of chlorella, therefore can be to chlorella
Culture production bring about great losses.At present, in chlorella culture the method for miscellaneous bacteria is controlled mainly to have Physical and chemical method, object
Logos mainly includes centrifugation and filtering, and chemical method mainly includes addition fungicide and antibiotic.Either physics or chemistry side
Method causes using effect unobvious, fungicide and antibiotic to pollute, and environment and influence frustule albumen quality etc. are a series of to ask
Topic, so far chlorella culture germ contamination are the sections for restricting chlorella industrialization production there has been no a kind of good solution
Grind problem.
The content of the invention
The present invention seeks to be directed to the problem of restriction chlorella industrialization production, a kind of utilization bacteriophagic Bdellovibrio is provided and is controlled
The method of germ contamination in chlorella culture.This method is small to control by adding bacteriophagic Bdellovibrio in chlorella incubation
The quantity of other bacteriums in the culture of ball algae, so as to improve the growth of chlorella.This method performance is stablized, and can be applied to industry
Change.
For this purpose, the present invention provides it is a kind of using bacteriophagic Bdellovibrio control chlorella culture in germ contamination method,
By the way that bacteriophagic Bdellovibrio is added in chlorella cultivating system, to control and/or reduce other bacterial numbers, and then promote small
The growth of ball algae.
In the present invention, the bacteriophagic Bdellovibrio include Bdellovibrio in it is all possess infect kill gramnegative bacterium and
One or more of Bdellovibrio of part gram-positive bacterium.
In the present invention, the chlorella includes one or more of all chlorellas in Chlorella.
According to certain embodiments of the present invention, the described method includes bacteriophagic Bdellovibrio cell suspension is added to bead
The step of being cultivated in algae cultivating system, wherein, the bacteriophagic Bdellovibrio cell suspension is by infecting Gram-negative bar
The culture of bacterium obtains.
In some preferred embodiments of the present invention, the corresponding bacterial strain of the bacteriophagic Bdellovibrio is that deposit number is
The bacteriophagic Bdellovibrio bacterial strain of CGMCC NO.15141 or other any one or more bacteriophagic Bdellovibrio bacterial strains.
In some embodiments of the invention, with the gauge of chlorella cultivating system, the bacteriophagic Bdellovibrio cell just
Beginning concentration is 10/mL to 104A/mL.
In other embodiments of the present invention, the cell concentration of the bacteriophagic Bdellovibrio cell suspension is 107-8A/
mL。
According to certain embodiments of the present invention, the bacteriophagic Bdellovibrio cell suspension cultivates the starting stage in chlorella
It adds in or finds to add in the case of germ contamination in chlorella incubation.
In the present invention, the chlorella culture includes the illumination autotrophy culture of chlorella, mixed breeding culture and no light heterotrophism
It is one or more of in culture.
In some preferred embodiments of the present invention, the chlorella culture is the illumination autotrophy culture of chlorella.
In some further preferred embodiments of the present invention, the temperature of chlorella illumination autotrophy culture is 20-35 DEG C,
Further preferably 25-30 DEG C.
In other further preferred embodiments of the present invention, the light source for chlorella culture illumination is daylight
Lamp/or sunlight；It is preferred that the intensity of illumination of the illumination autotrophy culture is 2000-8000Lx.
In general different degrees of germ contamination can all occur in chlorella incubation, it is general using centrifugation at present
The physical methods such as filtering and the pollution level of addition fungicide or antibiotic etc. chemical methodes control bacterium, but the effect generated is not
It is apparent and a series of problems, such as frustule albumen quality is caused to decline, so far around chlorella culture germ contamination problem there has been no
A kind of good solution is the scientific research problem for restricting chlorella industrialization culture.
The method of germ contamination in the control chlorella culture provided by the present invention using bacteriophagic Bdellovibrio, by bead
Bacteriophagic Bdellovibrio is added in algae incubation, the quantity of other bacteriums in chlorella culture is successfully controlled, is greatly improved
The growth of chlorella has important application value.Pass through a series of different research experiments, it was demonstrated that this technical performance is steady
It is fixed, it can be applied to industrialization.
Description of the drawings
To be readily appreciated that the present invention, illustrate the present invention below in conjunction with the accompanying drawings.
Fig. 1 is the stream of bacteriophagic Bdellovibrio-YQ, bacterium and chlorella ellipsoidea cell distribution in chlorella culture of the invention
Formula cell instrument scatter diagram.
Fig. 2 shows that bacteriophagic Bdellovibrio-YQ cracks the growth curve of e. coli bl21 cell by infecting.
Fig. 3 shows the effect for by the bacteriophagic Bdellovibrio-YQ for adding different initial concentrations chlorella ellipsoidea being promoted to grow.
Fig. 4, which is shown, to be controlled by the bacteriophagic Bdellovibrio-YQ for adding different initial concentrations in chlorella ellipsoidea cultivating system
The effect of other bacterial numbers
Fig. 5 is shown to be bitten by the bacteriophagic Bdellovibrio-YQ for adding different initial concentrations in chlorella ellipsoidea cultivating system
The situation of bacterium Bdellovibrio-YQ growths.
Culture presevation
Bacteriophagic Bdellovibrio (Bdellovibrio bacteriovorus), by the biological development corporation, Ltd. point in Guangdong more than green hundred
From, identification, China Committee for Culture Collection of Microorganisms's common micro-organisms center (referred to as：CGMCC；Address：Beijing
No. 3 Institute of Microorganism, Academia Sinica of institute of Chaoyang District North Star West Road 1) preservation, preservation date：On January 2nd, 2018, preservation
Number：CGMCC NO.15141.The bacterial strain is being named as-YQ plants of (Bdellovibrio of bacteriophagic Bdellovibrio in the present invention
bacteriovorus-YQ)。
Specific embodiment
To make the present invention easier to understand, below in conjunction with drawings and examples, the present invention will be described in detail, these realities
Apply example only serve it is illustrative, it is not limited to application range of the invention, unmentioned specific experiment side in the following example
Method is usually carried out according to routine experiment method.
As previously mentioned, different degrees of germ contamination can all occur in chlorella incubation, either using physics
Miscellaneous bacteria in method or chemical method control chlorella culture causes using effect unobvious, fungicide and antibiotic pollution
A series of problems, such as environment and influence frustule albumen quality, therefore there has been no one for germ contamination problem in chlorella culture so far
The good solution of kind, becomes the scientific research problem for restricting chlorella industrialization production.
To seek the new way of germ contamination in efficiently removal chlorella culture, the present inventor is in chlorella incubation
Germ contamination problem carried out substantial amounts of research.The present inventor passes through unremitting research and probe, finally from prawn culturing bottom of pond
One plant has successfully been filtered out in mud and has possessed the microorganism fungus kind for efficiently killing ability to the polluted bacteria in chlorella culture, through dividing
From-YQ plants of bacteriophagic Bdellovibrio (Bdellovibrio bacteriovorus-YQ) is accredited as, testing result shows the bacterial strain pair
Polluted bacteria in chlorella culture Bacteria Culture possesses efficient killing ability；The present inventor further study show that, to bead
Algae cultivating system addition bacteriophagic Bdellovibrio can kill the bacterium of pollution, effectively control and phagocytosis leech arc is removed in chlorella cultivating system
The quantity of bacterium beyond bacterium-YQ, and then improve the growth of chlorella.The present invention is based on what above-mentioned discovery was made.
Therefore, the side using germ contamination in bacteriophagic Bdellovibrio control chlorella culture involved by first aspect present invention
Method, be by the way that bacteriophagic Bdellovibrio is added in chlorella cultivating system, to control and/or reduce other polluted bacteria quantity,
And then promote the growth of chlorella.
In the present invention, the bacteriophagic Bdellovibrio include Bdellovibrio in it is all possess infect kill gramnegative bacterium and
One or more of Bdellovibrio of part gram-positive bacterium.
In the present invention, the chlorella includes one or more of all chlorellas in Chlorella；It is preferred that chlorella
For chlorella ellipsoidea.For example, chlorella ellipsoidea (Chlorella ellipsoide, number FACHB- used in the present invention
41) purchase is in Inst. of Hydrobiology, Chinese Academy of Sciences.The Escherichia coli germline used generally uses in genetic engineering operation
And the e. coli bl21 (E.coli BL21) of no pathogenicity.
Bacteriophagic Bdellovibrio (Bdellovibrio bacteriovorus sp.) is that one kind can prey on other all leather orchids
Family name's negative bacillus and the gramnegative bacterium of part gram-positive bacterium, foreign scholar in 1962 from pseudomonad for the first time
It was found that Bdellovibrio, hereafter finds the presence for having Bdellovibrio in soil, water body and animal body and people's enteron aisle.Bdellovibrio is basis
The name of its fundamental characteristics, mono- words of Bdeollvibrio derive from Latin language, and bdella is the meaning of " leech ", and vibrio is
The meaning of " arcuation " connects together as parasitic arcuation bacterium.
Bacteriophagic Bdellovibrio (Bdellovibrio bacteriovorus sp.) mainly by infect Gram-negative bacteria and
Part gram-positive bacteria is bred, and not yet finds that bacteriophagic Bdellovibrio infects the Eukaryotic report such as chlorella so far,
It is therefore contemplated that it does not generate any harm to the growth of chlorella.Moreover, inventor also demonstrates phagocytosis leech by experiment
Vibrios will not infect chlorella, and any harm is not generated to the growth of chlorella.And allow people surprisingly, inventor's warp
It crosses experiment to find, bacteriophagic Bdellovibrio can also be used as chlorella growth by cracking the nutriment discharged after other bacterial cells
Nutriment, and then more promote the growth of chlorella, there is important application value in chlorella industrialization culture.
In some embodiments of the invention, the thin of pollution can be killed to chlorella cultivating system addition bacteriophagic Bdellovibrio
Bacterium effectively controls other bacterial numbers in chlorella cultivating system in addition to bacteriophagic Bdellovibrio, and then is greatly improved small
The growth of ball algae.
According to certain embodiments of the present invention, the described method includes bacteriophagic Bdellovibrio cell suspension is added to bead
The step of being cultivated in algae cultivating system, wherein, the bacteriophagic Bdellovibrio cell suspension is by infecting Gram-negative bar
The culture of bacterium obtains.
In some preferred embodiments of the present invention, the corresponding bacterial strain of the bacteriophagic Bdellovibrio is that deposit number is
Bacteriophagic Bdellovibrio-YQ the bacterial strains of CGMCC NO.15141 or other any one or more bacteriophagic Bdellovibrio bacterial strains.
In other embodiments of the present invention, the cell concentration of the bacteriophagic Bdellovibrio cell suspension is 107-8A/
mL。
The present inventor employs flow cytometer (CY-S-3001, German Partec first；Major parameter is arranged to：FSC
155, SSC 290, FL1 248, FL2 361.5 selects the blue laser that exciting light is 355nm), the significance difference according to cell size
It is different, successfully bacteriophagic Bdellovibrio, the distribution of bacterium and chlorella cells and the quantity in chlorella culture are carried out for the first time
Identification and counting, for example, the flow cytometer scatterplot that bacteriophagic Bdellovibrio, bacterium and chlorella cells are distributed in chlorella culture
Figure is as shown in Figure 1.
In the present invention, bacteriophagic Bdellovibrio-YQ used is obtained in the following ways：
In clean bench under aseptic condition, cell is prepared using normal saline solution of 500 milliliters of triangular flasks in sterilizing
Concentration 107100 milliliters of the e. coli bl21 cell suspending liquid of a/mL is inoculated with initial cell density 2.6 × 103A/mL's bites
After bacterium Bdellovibrio-YQ, cultivated 5 days under the conditions of 30 DEG C of 200 revs/min of shaking speeds, by flow cytometer to Escherichia coli
BL21 cells and bacteriophagic Bdellovibrio-YQ cell concentrations variation curve show (as shown in Figure 2), cultivate 5 days after bacteriophagic Bdellovibrio-
The cell concentration of YQ increases to 6.3 × 107A/mL.By bacteriophagic Bdellovibrio-YQ (cell containing e. coli bl21) culture mistake
After 0.45 μm of filter membrane, size can be retained and be 1 μm or more of e. coli bl21 cell, and size is contained only in filtrate as 0.2 μ
Bacteriophagic Bdellovibrio-YQ the cells of m or so thus obtain the phagocytosis leech controlled in being cultivated for chlorella used in germ contamination
Vibrios-YQ cell suspending liquids, cell concentration have reached 107-8Between a/mL.
In some specific embodiments of the present invention, the preparation method of the bacteriophagic Bdellovibrio cell suspension includes：
(1) prepare sterilizing LB fluid nutrient mediums (tryptone 10g, yeast extract 5g, NaCl 10g, water 1000mL,
PH7.2), it is inoculated with e. coli bl21 (E.coli BL21) and carries out shaking table shaken cultivation, 200 revs/min of rotating speed, temperature 37 afterwards
℃.Culture centrifuges (10000 revs/min, 10 minutes) harvest Bacillus coli cells after 1 day, utilize the lifes through 121 DEG C of sterilizings in 10 minutes
Saline solution (0.9% sodium chloride) is managed, is centrifuged again after suspension Bacillus coli cells, Escherichia coli are obtained after repeated washing 3 times
Cell suspending liquid, for cultivating bacteriophagic Bdellovibrio.
(2) after being inoculated with bacteriophagic Bdellovibrio in Bacillus coli cells suspension, under 30 DEG C of 200 revs/min of shaking speeds
It is cultivated, the variation of Bacillus coli cells and bacteriophagic Bdellovibrio cell concentration, Fig. 2 display inoculations is measured by flow cytometer
After bacteriophagic Bdellovibrio-YQ cultures, Bacillus coli cells concentration continues to decline, and bacteriophagic Bdellovibrio cell concentration persistently rises, and says
Bright bacteriophagic Bdellovibrio possesses the ability for infecting typical gram-Negative bacillus Escherichia coli.By bacteriophagic Bdellovibrio (bar containing large intestine
Bacterium cell) after culture crosses 0.45 μm of filter membrane, the Bacillus coli cells that size is 1 μm or more can be retained, and in filtrate containing only
The bacteriophagic Bdellovibrio cell for having size to be 0.2 μm or so thus obtains and germ contamination institute is controlled in being cultivated for chlorella
Bacteriophagic Bdellovibrio cell suspending liquid, cell concentration have reached 107-8A/mL.
The training method of chlorella is not particularly limited in the present invention, for example, it is small that following methods culture may be employed
Ball algae：Sterilizing bead algae culture medium, inoculation chlorella ellipsoidea (Chlorella ellipsoide, number are prepared according to table 1
FACHB-41 after) intensity of illumination be 6000lx, illumination/interlunation be 16h/8h, temperature be 27 DEG C at carry out illumination autotrophy
Culture.It in general all can be inevitable there are different degrees of germ contamination, if be not controlled by chlorella incubation
Bacterial number is caused to increase and influence the growth of chlorella.
1 chlorella culture medium prescription of table
In the present invention, the bacteriophagic Bdellovibrio cell suspension adds in or in the chlorella culture starting stage in bead
It finds to add in the case of germ contamination in algae incubation.Such as：In some embodiments of the invention, cultivated in chlorella
In the process, it is inoculated with to infect the bacteriophagic Bdellovibrio for cracking bacterium while chlorella is inoculated with, so can successfully controls small
The growth of other bacteriums in ball algae cultivating system, and then promote the growth of chlorella.Bead in chlorella incubation
Frustule, bacterial cell and the variation of bacteriophagic Bdellovibrio cell concentration are identified and are measured using flow cytometer.
In chlorella incubation the phagocytosis leech arc of different initial cell densities can be obtained by being inoculated with different amounts
Bacterium-YQ.For example, in some embodiments, the phagocytosis of different initial cell densities is with the addition of in chlorella incubation respectively
Bdellovibrio-YQ, by continuously detecting chlorella ellipsoidea, the variation of bacterium and bacteriophagic Bdellovibrio-YQ cell concentrations, it was demonstrated that pass through
Addition bacteriophagic Bdellovibrio can successfully control the quantity of other polluted bacterias, while significantly promote the life of chlorella ellipsoidea
It is long.It is found by a series of experiments, this biological prevention and control high-tech is stable and reliable for performance, can be used for the high density training of chlorella
It supports.
In some preferred embodiments of the present invention, in terms of the culture solution in chlorella cultivating system, the phagocytosis leech
The initial concentration of vibrios cell is 10/mL to 104A/mL.
In the present invention, the chlorella culture includes the illumination autotrophy culture of chlorella, mixed breeding culture and no light heterotrophism
It is one or more of in culture.
In some preferred embodiments of the present invention, the chlorella culture is the illumination autotrophy culture of chlorella.
In some further preferred embodiments of the present invention, the temperature of chlorella illumination autotrophy culture is 20-35 DEG C,
Further preferably 25-30 DEG C.
In other further preferred embodiments of the present invention, the light source for chlorella culture illumination is daylight
Lamp/or sunlight；It is preferred that the intensity of illumination of the illumination autotrophy culture is 2000-8000Lx.
The present invention is ultrapure in the case where not specifying for " water " in dilution or culture medium process for preparation
Water.
Term " chlorella cultivating system " of the present invention includes chlorella before bacteriophagic Bdellovibrio bacteria suspension is added in and trains
Base and chlorella are supported, optional further includes other polluted bacterias；It adds after bacteriophagic Bdellovibrio bacteria suspension including chlorella
Culture medium and chlorella, bacteriophagic Bdellovibrio bacteria suspension (normal saline solution+bacteriophagic Bdellovibrio), optional further includes other pollutions
Bacterium.
Term " polluted bacteria " of the present invention or " other polluted bacterias " refer to influence bead in chlorella cultivating system
The bacterium in addition to bacteriophagic Bdellovibrio of algae growth, is mainly Gram-negative bacteria.
Heretofore described term " initial concentration of the bacteriophagic Bdellovibrio cell " refers to total with chlorella cultivating system
The number of the bacteriophagic Bdellovibrio cell of (volume) meter is measured, unit is " a/mL ".
Bacteriophagic Bdellovibrio-YQ, bacterium and chlorella ellipsoidea cell concentration are measured using following methods in the present invention：
The measure of bacteriophagic Bdellovibrio-YQ, bacterium and chlorella ellipsoidea cell concentration in chlorella ellipsoidea liquid culture,
Liquid culture is taken, after normal saline solution dilutes certain multiple, directly using flow cytometer (CY-S-3001, Germany
Partec；Major parameter is arranged to：FSC 155, SSC 290, FL1 248, FL2 361.5 selects the indigo plant that exciting light is 355nm
Laser) measure different cell concentrations therein.
Embodiment
Embodiment 1：Bacteriophagic Bdellovibrio-YQ is not added in chlorella ellipsoidea cultivating system
1. the preparation of bacteriophagic Bdellovibrio-YQ：
100mL normal saline solutions (0.9% sodium chloride) are prepared in 500 milliliters of triangular flasks, through 121 DEG C of sterilizings of high temperature and pressure
It is put into after twenty minutes in clean bench and carries out ultraviolet disinfection 10 minutes.In clean bench under aseptic condition, 1mL is added in
Cell concentration is 1011The e. coli bl21 cell suspension of a/mL, while picking is seeded in the phagocytosis of growth in double-layer plate
Bdellovibrio-YQ carries out culture 5 days at 30 DEG C of temperature, 200 rpms of shaking speed.In clean bench, culture is taken
Cell concentration can be obtained by, which crossing after 0.45 μm of filter membrane, reaches 6.3 × 107A/mL bacteriophagic Bdellovibrio-YQ cell suspensions.
2. the culture of chlorella ellipsoidea：Culture medium is prepared according to 1 chlorella culture medium prescription of table, in 500 milliliters of triangular flasks
100 milliliters of culture mediums of people are inside put, is put into after twenty minutes in clean bench through 121 DEG C of sterilizings of high temperature and pressure and carries out ultraviolet disinfection
10 minutes.In clean bench under aseptic condition, initial frustule concentration is 6.0 after adding in 1mL chlorella ellipsoidea cultures
×106A/mL.500 milliliters of triangular flasks for being inoculated with chlorella ellipsoidea are put into illumination box, are in intensity of illumination
6000lx, illumination/interlunation are 16h/8h, and temperature is that illumination autotrophy culture is carried out at 27 DEG C.
3. chlorella and the growth of bacterium
It is not added in chlorella incubation under the conditions of bacteriophagic Bdellovibrio-YQ, chlorella ellipsoidea cell is dense after cultivating 5 days
It spends from 6.0 × 106A/mL increases to 2.3 × 107A/mL (Fig. 3), at the same time polluted bacteria is thin in chlorella ellipsoidea culture
Born of the same parents' concentration is from initial 5.6 × 106A/mL increases to 1.9 × 107A/mL (Fig. 4).This result illustrates in chlorella incubation
In, with the growth of chlorella, polluted bacteria is also in further growth.
Embodiment 2：It is 10 that initial cell density is added in chlorella ellipsoidea cultivating system3Bacteriophagic Bdellovibrio-the YQ of a/mL
Bacteriophagic Bdellovibrio-YQ and chlorella ellipsoidea culture are prepared same as Example 1 in embodiment 2
It is removed in embodiment 2 and initial cell density is with the addition of in chlorella ellipsoidea culture as 103The phagocytosis leech arc of a/mL
Bacterium-YQ is outer (1.6 μ L bacteriophagic Bdellovibrio-YQ cell suspensions are with the addition of in initial 100 milliliters of chlorella cultures), other controls
Condition is same as Example 1.
It is 10 that initial cell density is added in chlorella ellipsoidea incubation3Bacteriophagic Bdellovibrio-YQ the conditions of a/mL
Under, chlorella ellipsoidea cell concentration is from 6.0 × 10 after culture 5 days6A/mL increases to 3.2 × 107A/mL (Fig. 3), than not adding
39% algae biomass is improved during bacteriophagic Bdellovibrio-YQ.At the same time in chlorella ellipsoidea culture polluted bacteria concentration from
Initial 5.6 × 106A/mL is reduced to 2.1 × 105A/mL (Fig. 4), bacteriophagic Bdellovibrio-YQ cell concentrations are from 103A/mL increases
It is added to 4.8 × 107A/mL (Fig. 5).This result illustrates that in chlorella incubation addition initial cell density is 103A/mL
Bacteriophagic Bdellovibrio-YQ, can not only reduce the polluted bacteria quantity in chlorella ellipsoidea cultivating system, but also can promote
The growth of chlorella ellipsoidea.
Embodiment 3：It is 10 that initial cell density is added in chlorella ellipsoidea cultivating system4Bacteriophagic Bdellovibrio-the YQ of a/mL
Bacteriophagic Bdellovibrio-YQ and chlorella ellipsoidea culture are prepared same as Example 1 in embodiment 3.
It is removed in embodiment 3 and initial cell density is with the addition of in chlorella ellipsoidea culture as 104The phagocytosis leech arc of a/mL
Bacterium-YQ is outer (3.2 μ L bacteriophagic Bdellovibrio-YQ cell suspensions are with the addition of in initial 100 milliliters of chlorella cultures), other controls
Condition is same as Example 1.
It is 10 that initial cell density is added in chlorella ellipsoidea incubation4Bacteriophagic Bdellovibrio-YQ the conditions of a/mL
Under, chlorella ellipsoidea cell concentration is from 6.0 × 10 after culture 5 days6A/mL increases to 3.5 × 107A/mL (Fig. 3), than not adding
52% algae biomass is improved during bacteriophagic Bdellovibrio-YQ.At the same time in chlorella ellipsoidea culture polluted bacteria concentration from
Initial 5.6 × 106A/mL is reduced to 5.5 × 104A/mL (Fig. 4), bacteriophagic Bdellovibrio-YQ cell concentrations are from 104A/mL increases
It is added to 1.6 × 108A/mL (Fig. 5).This result illustrates that in chlorella incubation addition initial cell density is 104A/mL
Bacteriophagic Bdellovibrio-YQ further reduce polluted bacteria quantity in chlorella cultivating system, and further promote ellipse
The growth of chlorella.
In conclusion the polluted bacteria in chlorella ellipsoidea incubation can be successfully controlled using bacteriophagic Bdellovibrio-YQ
Quantity, and then the growth of chlorella ellipsoidea is greatly improved.
The foregoing is merely the preferable realities for promoting chlorella growth using bacteriophagic Bdellovibrio control germ contamination of the present invention
Apply example, be not intended to limit the invention, within the spirit and principles of the invention, any modification for being made, equivalent substitution or
Improve etc., it should all be included in the protection scope of the present invention.
Claims (10)
1. a kind of method of germ contamination in control chlorella culture using bacteriophagic Bdellovibrio, by the way that bacteriophagic Bdellovibrio is added in
Into chlorella cultivating system, to control and/or reduce other polluted bacteria quantity, and then promote the growth of chlorella.
2. according to the method described in claim 1, it is characterized in that, the bacteriophagic Bdellovibrio possesses including all in Bdellovibrio
Infect one or more of Bdellovibrio for killing gramnegative bacterium and part gram-positive bacterium.
3. according to the method described in claim 1, it is characterized in that, the chlorella is included in Chlorella in all chlorellas
One or more.
4. according to the method described in any one in claim 1-3, which is characterized in that it includes hanging in bacteriophagic Bdellovibrio cell
Turbid is added to the step of being cultivated in chlorella cultivating system, wherein, the bacteriophagic Bdellovibrio cell suspension is by invading
The culture for contaminating gram-Negative bacillus obtains.
5. according to the method described in claim 4, it is characterized in that, the corresponding bacterial strain of the bacteriophagic Bdellovibrio is deposit number
For the bacteriophagic Bdellovibrio bacterial strain of CGMCC NO.15141 or other any one or more bacteriophagic Bdellovibrio bacterial strains.
6. method according to claim 4 or 5, which is characterized in that with the gauge of chlorella cultivating system, the phagocytosis leech
The initial concentration of vibrios cell is 10/mL to 104A/mL.
7. according to the method described in any one in claim 4-6, which is characterized in that the bacteriophagic Bdellovibrio cell suspension
Cell concentration be 107-8A/mL.
8. according to the method described in any one in claim 4-7, which is characterized in that bacteriophagic Bdellovibrio cell suspension is small
The ball algae culture starting stage adds in or finds to add in the case of germ contamination in chlorella incubation.
9. according to the method described in claim 1-8, which is characterized in that the chlorella culture includes the illumination autotrophy of chlorella
It is one or more of in culture, mixed breeding culture and no light Heterotrophic culture.
10. according to the method described in claim 9, it is characterized in that, chlorella culture is trained for the illumination autotrophy of chlorella
It supports；It is preferred that the temperature of chlorella illumination autotrophy culture is 20-35 DEG C, further preferably 25-30 DEG C；It is preferred for chlorella training
The light source for supporting illumination is fluorescent lamp/or sunlight；It is preferred that the intensity of illumination of the illumination autotrophy culture is 2000-8000Lx.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810030528.1A CN108102924B (en) | 2018-01-12 | 2018-01-12 | Method for controlling bacterial pollution in chlorella culture by using bdellovibrio bacteriovorus |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810030528.1A CN108102924B (en) | 2018-01-12 | 2018-01-12 | Method for controlling bacterial pollution in chlorella culture by using bdellovibrio bacteriovorus |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN108102924A true CN108102924A (en) | 2018-06-01 |
| CN108102924B CN108102924B (en) | 2021-06-25 |
Family
ID=62220016
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810030528.1A Active CN108102924B (en) | 2018-01-12 | 2018-01-12 | Method for controlling bacterial pollution in chlorella culture by using bdellovibrio bacteriovorus |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108102924B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114164128A (en) * | 2021-12-15 | 2022-03-11 | 海南绿藻世界生物科技有限公司 | Algae symbiotic bacteria composition, algae bacteria co-culture system and microalgae culture method |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20110166844A1 (en) * | 2003-08-01 | 2011-07-07 | Dna Twopointo, Inc | Systems and Methods for Antibody Engineering |
| CN105016486A (en) * | 2015-07-27 | 2015-11-04 | 莆田市天然星农业开发有限公司 | Control formula and enforcement method aiming at enteromorpha harm during polyculture of shrimp, crab and shellfish |
| CN106396130A (en) * | 2016-11-16 | 2017-02-15 | 防城港市鑫润养殖有限公司 | Prevention and control method for blue-green algae in marine shrimp culture water |
-
2018
- 2018-01-12 CN CN201810030528.1A patent/CN108102924B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20110166844A1 (en) * | 2003-08-01 | 2011-07-07 | Dna Twopointo, Inc | Systems and Methods for Antibody Engineering |
| CN105016486A (en) * | 2015-07-27 | 2015-11-04 | 莆田市天然星农业开发有限公司 | Control formula and enforcement method aiming at enteromorpha harm during polyculture of shrimp, crab and shellfish |
| CN106396130A (en) * | 2016-11-16 | 2017-02-15 | 防城港市鑫润养殖有限公司 | Prevention and control method for blue-green algae in marine shrimp culture water |
Non-Patent Citations (1)
| Title |
|---|
| 刘春娇: "蛭弧菌在球等鞭金藻培养中的应用研究", 《中国优秀硕士学位论文全文数据库 农业科技辑》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114164128A (en) * | 2021-12-15 | 2022-03-11 | 海南绿藻世界生物科技有限公司 | Algae symbiotic bacteria composition, algae bacteria co-culture system and microalgae culture method |
Also Published As
| Publication number | Publication date |
|---|---|
| CN108102924B (en) | 2021-06-25 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Creswell | Phytoplankton culture for aquaculture feed. | |
| CN109097298B (en) | Method for preparing bdellovibrio bacteriovorus preparation by enrichment culture method | |
| US6673592B1 (en) | Continuous cultivation of microorganisms in large open tanks in sunlight | |
| CN104611227B (en) | Scenedesmus obliquus with tolerance to high pH and breeding method thereof | |
| CN103773687B (en) | A kind of cultural method of microalgae | |
| CN110184222A (en) | A kind of bacteriophagic Bdellovibrio freeze-dried powder preparation and its application | |
| CN110157621B (en) | Preparation method of high-concentration microalgae living cell long-acting preservative | |
| CN104312963A (en) | Method for separating purified bdellovibrio from active sludge | |
| Muskat et al. | Fermentation of the psyllid-pathogenic fungus Pandora sp. nov. inedit.(Entomophthorales: Entomophthoraceae) | |
| CN105838683A (en) | Method for proliferation of mink canine distemper virus by applying novel cell microcarrier | |
| CN108102924A (en) | A kind of method of germ contamination in control chlorella culture using bacteriophagic Bdellovibrio | |
| CN106190898B (en) | Industrial liquid fermentation method of bacillus cereus JZBC1 for dissolving pond dinoflagellate | |
| CN104974949B (en) | A kind of screening technique for seawater fish shield infusorian cause of disease insecticidal bacteria | |
| CN106701645A (en) | Bacillus amyloliquefaciens B7 with immune growth promoting effect and application method of bacillus amyloliquefaciens B7 | |
| CN109609405A (en) | Produce bacillus and the purposes of algistatic activity substance | |
| KR101797070B1 (en) | Culture compositions for promoting growth of spirulina and the method culturing for promoting growth of spirulina by using the same | |
| CN104845940B (en) | Salmonella phage preparation and preparation method thereof | |
| CN103146597A (en) | Method for preparing photosynthetic bacteria liquid | |
| CN106635865A (en) | Culture medium for separation and purification of mycoplasma bovirhinis, preparation method and application of culture medium | |
| CN107058212A (en) | Can Secondary Culture Testis Caprae seu Ovis passage cell and its passage and attenuation cultural method and special cultivating system and application | |
| CN106967610B (en) | Culture method of spherical phaeocystis | |
| Pan | Growth of a photoautotroph, Plectonema boryanum, in the dark on glucose | |
| CN105176887A (en) | Method for culturing escherichia coli | |
| CN113100129B (en) | Culture medium and culture method for long-term and continuous culture of back mouth insects | |
| CN110257250A (en) | The culture media composition and cultural method and bioreactor of Chaetoceros muelleri |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CP02 | Change in the address of a patent holder |
Address after: 524051 No.8, Yexing Road, D District, Guandu Industrial Park, Potou District, Zhanjiang City, Guangdong Province Patentee after: GUANGZHOU LYUBAIDUO BIOLOGICAL DEVELOPMENT Co.,Ltd. Address before: 524051 No.8, Yexing Road, D District, Guandu Industrial Park, Guandu town, Potou District, Zhanjiang City, Guangdong Province Patentee before: GUANGZHOU LYUBAIDUO BIOLOGICAL DEVELOPMENT Co.,Ltd. |
|
| CP02 | Change in the address of a patent holder | ||
| PE01 | Entry into force of the registration of the contract for pledge of patent right |
Denomination of invention: A method for controlling bacterial pollution in Chlorella culture by using bacteriophage Bdellovibrio Effective date of registration: 20220510 Granted publication date: 20210625 Pledgee: Zhanjiang Branch of China Construction Bank Co.,Ltd. Pledgor: GUANGZHOU LYUBAIDUO BIOLOGICAL DEVELOPMENT Co.,Ltd. Registration number: Y2022980005307 |
|
| PE01 | Entry into force of the registration of the contract for pledge of patent right |